Introduction of fecal bacteria into germ-free (GF) Balb/c mice induces class II major histocompatibility complex (MHC) molecules on the small intestinal epithelium. In this study, we elucidated the regulatory mechanisms for the class II MHC molecule induction on the mouse small intestinal epithelium during microbial colonisation of the gut in ex-GF mice. Intraperitoneal injection of interferon-gamma (IFN-gamma) into GF Balb/c mice induced class II MHC expression on the small intestinal epithelial cells. Induction of these molecules was inhibited by peritoneal injection of a monoclonal antibody (mAb) against IFN-gamma on the conventionalisation of GF mice. RNA reverse transcriptase-polymerase chain reaction (RT-PCR) analysis of the small intestinal epithelium indicated that the class II transactivator (CIITA), a regulatory factor for the class II MHC gene, and the IE alpha chain, but not IFN-gamma receptor mRNA, increased during conventionalisation. The induction of class II MHC on the epithelial cells during the conventionalisation of GF CB-17 scid mice was much lower than that in GF Balb/c mice. Immunocytochemical and RT-PCR analysis showed that both the number of IFN-gamma producing IEL and the level of the IFN-gamma mRNA in gamma delta TCR IEL were very low in the GF state, and gradually increased after microbial colonisation. After in vivo treatment with a mAb against gamma delta TCR, the number of gamma delta TCR-expressing IEL greatly decreased and the expression of class II MHC molecules on the small intestinal epithelium was repressed during the conventionalisation of GF mice. Taken together, these results suggested that gamma delta TCR-bearing IEL modulate class II MHC molecule expression on the small intestinal epithelium through the production of IFN-gamma during microbial colonisation in ex-GF mice.